Therapeutic colloid



Patented Apr. 18 1939 PATENT OFFICE THERAPEUTIC COLLOID Garth WilkinsonBoericke and William Wallace Young, Philadelphia, Pa., assignors to J.C. Shay, Incorporated, Philadelphia, Pa.

No Drawing. Application April 27, 1936, Serial No. 76,602

4 Claims.

Our invention relates to the art of the production of substances incolloidal state to be used therapeutically and more particularly to theproduction of fatty sols that adsorb the toxins that 5 pervade the humanblood stream when the body is in a state of unhealth.

The object of our invention is the preparation of stable fatty sols by anovel process which can be administered intravenously or subcutaneously10 for the purpose of adsorbing and fixing the toxins that arefrequently the evidentiary manifestation of disease and for the ultimatepurpose of restoring the health of an individual to an accepted.

- I which the name Blooddust had been generally applied. One of theearly observers had previously suggested the term Chylomicron for thesefine particles, inasmuch as investigation had established their fattycharacter and approximate Jo size, one-half to one micron; other factsestablished by the early workers were that chylomicrons had their originfrom fat and not from protein or carbohydrates, that the numbercirculating on the blood stream has a constant relaj tion tothe foodintake and the approximate nu- V merical relationship to other formedelements were estimated, i. e. in a cubic millimeter of blood there areabout eighty million chylomicrons to five thousand White and fivemillion red. The m last point seems to us specially important,thetremendous'quantity of these particles in relation to the blood cells.It is obvious that such a predominance of one subject in the blood serumshould have more biological significance than 5 simply the function ofnutrition and hence might logically be affected by the various states ofhealth and disease in the same manner that the red or white cells are. 1

Search of the literature for further .of this important emulsion broughtto light a series of papers by certain workers whose careful workconcerned itself with the physical and chemical aspects of the'chylomicron emulsion. They concluded that each fatty particle was llcoated with-a protein film, presumably a mixture knowledge ol albumenand globulin and further that the number of the circulating particleshad no effect on the specific gravity or the viscosity of the blood asshown by the methods they employed in their work.

Further research revealed that the extension ultramicroscopicinvestigation of the blood serum has been done by certain otherindividuals. These workers consider the electrophysical state of theparticles to be the determining factor in health and disease andutilizing the modern conception of the structure of the atom they feelthat an explanation of all diseased processes is possible by theirhypothesis. The essential fatty character of these minute particles isnot recognized by these last mentioned individuals but then, as has beenshown previously, the surface film may well be protein, this being amajor factor in the stabilization of such an emulsion.

From this work, there is no doubt that there is a very suggestiverelationship between the state of health of the individual and themorphology and characteristics of the small particles.

We have examined over a thousand cases under the dark field covering awide range of sickness and it is quite possible to describe an illnessin terms familiar to the physical chemist by observing theultra-microscopic picture of a blood serum.

Before discussing the common changes observed under the dark field, afew definitions should be in order:

Brownian movement-This is a vibratory motion peculiar to small particlesin a state of colloidal suspension. It is due apparently to theelectronic bombardment of the particles by the external phase.

Hydration.-Where the field shows a number of variations under differentheads, for examplea fewer number of particles-an increased size of theparticle-a sluggish repulsion between particlesa sluggish Brownianmovement-the presence of clumps.

Dispersion is the opposite of the above and is identified generally withthe normal appearing field-that is a goodly number of particles,excellent motion, usually uniform in size, and no clumps.

After stating the possible biological variations in the characteristicsof a fatty emulsion, we must note some of the more obvious changes,commonly observed by looking under the dark field. The commonestfindings are a lessened activity and diminution in the number of thesecirculating chylomicrons, especially in wasting diseases. In diseases ofan endotoxic character, clumping and agglutination under the miscroscopeof these individual particles seem to be a sign of infection or ofexisting toxemia. This clumping and agglutination by the fat particlesin the presence of a circulating toxin suggests that perhaps fat plays apart in detoxification. In other words, fats are toxiphylic and fix atoxin, the union being chemical or physical, and resultinginan innocuoussubstances.

The ability of a lipoid and fat to fix toxic radicals is well known, andcertainly the colloidal form circulating in the blood is, by virtue ofits tremendous surface development, singularly well adapted to thisrole.

Some examples of this power of fat toabsorb may be given: the aflinityof diphtheria and tetanus for the lipoids of our body. If wedelay ingiving antitoxin for instance, there will be no unbound toxinscirculating to meet it and, hence, the danger of delay in givingantitoxin is serious.

Again, We know that a prolonged toxemia of an organ leaves a depositionof fat in it. Fatty degeneration and infiltration are every dayexperiences to the pathologist. A kidney which has lost its functionthrough inflammation is apt to'be full of fat, a liver likewise reactsby having fatty replacements, especially in severe poisoning. It wasformerly believed that this depended on a'transformation of-the'proteinsof 'the tissues into fat, under the influence of the poison. It isevident that if such a change were as wide-spread asthe fat depositswould lead chematous cells is not formed from their own protein, but isdeposited after transportation from fat deposits already existing.

"Our point about this whole matter is-the assumption that a toxic organhas 'fat brought to'it by'nature in an effort to detoxicate the existingcondition and is not'simply a phonon enon'per se of the action of toxinon cells.

It'may not be an exogenous type like diphtheria or tetanus. It may beendogenous'orintracelldlanexerting its eifects only upon celldestruction. 'It may be on the other hand an untoward byeproduct ofderanged metabolism, or finally and most probably, it may be a so-calledchange of physical or chemical equilibrium and not'a separate substanceat all.

We have prepared lecithin sols but consider lanolin superior, which is atri-cholesterol-ester of'the three fatty acids palmitic, stearic andoleic. 'It is produced by condensation methodsusing alcohol and water,and when finished is extraordinarily stable, but extremely sensitive toa toxic radical. The actual method of procedure is indicated as follows:

Anhydrous lanolin is used because itis slightly soluble in alcohol andpartly mixes with water without separating.

Quantitative measures are not necessary in the preparation of this solbecause there is only a certain amount absorbed by the alcohol, therapidity and quantity varyin with temperature of'the alcohol. For thisreason warm alcohol is used, to save time and conserve alcohol, thoughitmay be made with cold. A. sufficient amount,

movements. 'ingthe gen'eralrule that the phase'having the lowerdielectric constant carries a negative charge, the positive phase inthis case being group turned toward the water.

usually about fifty grams of lanolin is stirred in a large beaker bymeans of a mechanical stirrer for one and a half to two hours. Samplesare taken after the first hour and mixed with an equal quantity ofdistilled water. If a prompt, white precipitate forms, which alterseveral samplesdoes not increase in density, this is taken as evidencedsufficient saturation.

The volume of lanolin-alcohol mixture is then mixed with three times itsquantity of distilled water and an immediate white emulsion is formed.

The next step is the driving ofi of the alcohol over a steam bath at atemperature varying from eighty to ninety degrees. The volume at the endof this process is roughly half that of the original 'mixture andalcohol should not be detected by smell although there is a small quan-'l'iave shown it to be sterile from all types of organism and'sporeformers. Freezing promptly breaks the emulsion, and agitation fails torestore it.

It may be stored in a cool place with acork 3 or cotton plug. Beforeclinical use, it is brought to a boil or is autoclaved,-and injectedslowly at body temperature.

Its'physical properties are: a milky-appearing aqueous'liquid,consisting of particles about one micron in size showing active BrownianIt carries a negative charge obeywater. its pH varies slightly, but maybe adjusted easily to 5.5. There is undoubtedly an orientation of thefatty molecules'with the polar Hence, any toxin with which it unitedmust present a similar active or polar group as recognized by colloidchemistry.

We contend that our emulsion is not stabilized by any film, dispersionbeing quite permanent even after three years observation. its stabilitydepends upon two factors: one being-the electric charge and the other ahydration factor-spoken of by Freundlich.

The sol is thermo-stable even at boiling temperature and ultra-violetlight rather increases the energy of the Brownian movement.

In a general way, dispersion is enhanced "with an increase of thepH or ashift to the basic side of the solution, While hydration is broughtabout by the addition of acids. 100 cc. of colloid by 30 minimum lethaldoses of diphtheria toxin'will occur inlanolin sol at a pH value of 4.5,but will not occur atapI-I value of 5.0. Most diseases of a toxic natureare associated generally with acidosis which is compensated for by thebuffer salts. In. these diseases,.we note routinely a state of markedhydration of the chylomicrons in the blood serum. Conversely in diseasescharacterized by alkalosis,fthe chylomicrons in the serum are routinelyhighly dispersed.

Presumably,

Thus,.precipitation of The effect of a number of substances upon thiscolloid was investigated next. From test-tube experiments, wedemonstrate that: a drop of toxic blood will precipitateapproximatelyacc. of this lanolin. Toxic blood is meant simply that the bloodcomes-from a case showing ordinary-symptoms of toxemia, such as fever,leucocytosis, malaise, sweat, etc.

Normal blood will notdo this. None of thefour blood typestaken from-awell person has any precipitating effect on the colloid. We have triedbecause it is the presumed afiinity of the fat for.-

toxin which causes a precipitation; and ifmany other substances didthis, it would be of no par ticular value. Other common chemicalsubstances do not do this. Thus methylene blue (10%) and phenol (97%),arsenic (1-100), mercurochrome (2%), bichloride ofmercury (10%) S. T,37, ceanothyn (tincture), iodine (sat. sol.), sodium hydroxide (16%)have no precipitating effect according to our experiments.

Other observations follow: Precipitation is caused regularly by apH'more acid than 1.2; but acidity is not the only factor as we testedmany sera which were definitely on the alkaline side, and a drop-ofwhich promptly precipitated the colloid. The promptness with whichcoagulation or precipitation takes place in the test tube after theaddition of a drop of toxic bloodfrom a sick patient is roughly parallelto the severity of the illness. Thus in a critcial pneumonia case in thefirst stage, precipitation was prompt in 4 minutes and when the test wasrepeated 1 week later, with the patient considerably bettenthe test took20 minutes before precipitation was noticeable. This phenomenon observedin vitro may be interpreted much as the blood sedimentation testfor redcells. 7

Of the various salts, the Cl ion seems to be particular in causingprecipitation in a lower concentration than other salts- Regarding thisprecipitation or agglutination phenomenon we assume that the fat fixes atoxic radical; and inour test-tube experiments this precipitating effectis considered evidence of toxicity. When acid is the precipitating agentthe result is due to'the H charge neutralizing the negative charge onthe particles.

Analyzing this phenomenon, it is clear that if the toxin in a toxicblood hasnot'been fixed by the emulsion in thetest-tube, the precipitate"if used again against another specimen ofcolloid in a test tube woulddrop it. But this will We found first that the colloid in 5 to 10 cc.doses harmful to humans.

was not harmful when injected into the earveins of a rabbit and doses of250 cc. are not The next step, we felt, was then to obtain a definitetoxin. of known strength instead of simply an unknown toxic serum. We

' obtained from a local laboratory diphtheria toxin represented thisdose.

otherseries of experiments, the conclusionsyof which follow:

Thefatal dosewhenmixed with-3 cc. of colloid andinjected together waswithout efiect. The control animal diedon thefourth day.

Three guinea pigs were given a lethal dose of" toxinv subcutaneously andthen, each was given respectively 1, 2, 3, cc. of colloid individed-doses starting 15 minutes after the original toxin injection andcontinuing every 2 hours for, 4 days. The results were that the guineapigs treated. bythe-colloids showed noeffects so far as We couldjudge bytheir appearance and behavior. The control animal died as before.

Further experiments brought out the fact that it takes more colloid toneutralize the toxin in the body.=than in the test-tube and also thatif, given all in one dose it is not as effective as the same amountdivided over a period.

Totest out the detoxicating power of fat in vivo, 3- minimum lethaldoses of toxin were put into 10cc. of colloid and allowed toprecipitate. At the end of 24 hours, the clear portion was injected intoav guinea pig and the precipitated portion was injected into anotherguinea pig. In both pigs no untoward effects were noted, whichwould'seem to indicate that the toxin had been adsorbedby theprecipitate rendering both clear portions and precipitate innocuous.Naturally, many experiments have been made with the end in view ofshaking loose the toxic portion after its union with the fat, all beingunsuccessful to date.

This however opensup a most fascinating investigation because, were itaccomplished, one would be in a position of being able to isolate anendogenous toxin, such as that of pneumonia, typhoid; grippe, etc., and.study it in its pure state as we do now'diphtheria, tetanus, andbotulinus.

In the preparation of this colloid for human injection, it was necessaryto find out whether or not the buffer salts have a precipitating effect.

The addition'of phosphates, sulphates and calciumcarbonate to thelanolin sol does not influence the state of dispersion, the Brownianmotion, the-sizeof the particles, or its therapeutic efficacy in any wayuntil a certain concentration is reached. Since this concentration iswell above that found physiologically, it is possible to employ a sol towhich has been added one or more of these salts in physiologicalproportions. The addition of sugar in the form of dextrose up to al0'per cent solution with or without the presence of a salt, such assodium carbonate, in no waymilitates against the stability of the sol.This finding is of value since it permits the use of a physiologicglucose solution (5 per cent).

Unadjusted, when freshly prepared, a colloidal sol of fat will be of acertain pH value. For instance, lanolin will have a pH value of about4.5 or a little higher. This is not the pH of the body, but from ourexperience gives excellent therapeutic results and even when used in asmuch as a 500 cc. dose has no deleterious effects. It is wise, however,and it is our custom to adjust our sols by the use of small quantitiesof a very dilute sodium hydroxide to: a pH of 7.5 or 7.6 before we usethem. It has been found that, if a sol of lanolin has a pH of 4.5, theaddition of sugar to make a 5 per cent solution will raise the pH valueto almost a physiological normal due to apparent depression ofionization. For this reason, it is hardly necessary to adjusta sol towhich glucose has been added. Somewhat the same effect, although not tothe same degree, follows the introduction of salt or sodium phosphate ina physiological percentage.

Our sols have routinely the appearance of milk and have a surfacetension slightly'above that of water. In each 1000 cc. there isapproximately 4.5 to 5.0 grams of fat. It is practically odorless andproduces nausea when an attempt is made to dring it.

Therapeutically, there is a choice among a number of preparations offat. We have used lanolin almost exclusively, although kekuni nut sol isjust as efiicacious. However, almost any other fat can be used. The solcan be used in its freshly-prepared state or with the addition ofdextrose. The method of administration depends upon the preparation tobe used. With the freshly-prepared sol, hypodermoclysis is the method ofchoice and as much as 250 cc. will be absorbed readily in minutes. Ifdextrose or the physiological salts or both are to be used, then theintravenous method is to be preferred, since the hypodermic method leadstodiscomfort because of the slow absorption. The intravenous use of thefreshly-prepared specimen will at times cause severe lumbar pains aftersome to 50 cc. have delivered. Since the material is kept as described,it is wise to boil the sol for 3 minutes before using, and to administerit at body temperature.

We have noted no sign nor symptoms of an allergic nature even of themost minute character in cases that we have treated. The initial dose isfrom 100 to 300 cc. depending upon the degree of toxicity. Thesubsequent doses, if necessary, are given at 2 to 5 day. intervals,depending upon the progress of the case. Where there is markeddehydration, anorexia, or imperfect nutrition, we find it wise to usethe sol containing sugar. Where the toxemic manifestations alonepredominate, we merely use the adjusted colloid. The indications for itsuse are the infectious diseased, from the mild cases exhibiting alow-grade toxicity which show merely a generalized aching and sorenessand tend to become chronic to the most virulent type of infections withtemperatures ranging as high as 107- degrees. Among the cases treatedsuccessfully the streptococcic and staphalicicin infections, cellulitis,septic arthritis, puerperal sepsis. Also we have observed the toxicsymptoms of tuberculosis improve after colloid injection (usuallysub-cutaneous).

Clinical toxic symptoms of the case serve as an indication, rather thanas a definite diagnosis. Also, since the affinity of the fat for thetoxin is not bacteriologically specific, the type of infecting organismis not of very great importance, and the colloid may be indicatedequally well in a case of typhoid, as in a caseof pneumonia ormeningitis. The best and most scientific indication for its use is to behad by a study of the serum under the dark field; and if hydration andclumps are marked, it is indicated.

Following the use of the lanolin with dextrose intravenously, there isfrequently met with a febrile reaction with a rise of temperature of 1or 2 degrees and a concomitant increase of the pulse rate. This isfollowed usually by a decrease of the temperature to normal orthereabouts. There may be in 24 or 48 hours another increase oftemperature, but it is wise to wait for at least '72 hours in a subacutecase, longer in a chronic case, or 24 hours in an acute case. Beforerepeating the dose, the indications for repetition are found in thebehavior of the temperature curve. The febrile reaction from a dose of200 cc. intravenously is marked by chills, fever, and sweat which lastfrom 15 to 20 minutes. This does not occur in all cases, however, andthe febrile reaction is avoided by the hypodermoclysis method ofadministration. Contra-indications have as yet not been met with,although the use of the colloid is apparently contra-indicated inathermatous subjects and in those under heavy digitalis dosage.

We are convinced that the colloid has no effect directly on bacteriathemselves, acting simply as a therapeutic sponge to absorb cellular andbacterial toxins and poisons and thereby give the patient a chance tocapitalize on his natural resistance or the other treatment instituted.

We are aware that the invention may be embodied in other specific formswithout departing from the spirit or essential attributes thereof, andwe therefore desire the present embodiment to be considered in allrespects as illustrative and nonrestrictive, reference being had to theappended claims rather than to the foregoing description to indicate thescope of our invention.

We claim:

1. A process for preparing lanolin sol which consists in agitatinglanolin with ethyl alcohol, mixing said lanolin and said ethyl alcoholwith approximately three times their volume of distilled water,expelling substantially all of said ethyl alcohol by heat, and filteringthe hydroalcoholic emulsion of said lanolin.

2. A detoxicant, therapeutic compound for parenteral use comprisinglanolin sol of colloidal particles of substantially 1 micron in size andunstabilized by other substances.

3. A detoxicant, therapeutic compound for subcutaneous use comprisinglanolin sol of colloidal particles of substantially 1 micron in size andunstabilized by other substances.

4. A detoxicant, therapeutic compound for intravenous administrationcomprising glucose and lanolin sol of colloidal particles ofsubstantially 1 micron in size and unstabilized by other substances.

GARTH WILKINSON BOERICKE. WILLIAM WALLACE YOUNG.

